This project focuses on the recognition of HLA-DR2/peptide complexes by myelin basic protein (MBP) specific T cell receptors (TCRs) that have been isolated from multiple sclerosis (MS) patients. It was previously thought that TCRs are highly specific for particular microbial or self-peptides. However, analysis of human MBP specific TCRs demonstrated a substantial degree of crossreactivity in TCR recognition of MHC-bound peptides. As a result, microbial peptides can activate these autoreactive T cell clones even though they only have limited sequence similarity with the self-peptide. The structural basis and biological significance of TCR crossreactivity will be examined in the proposed experiments. The goal of Aim 1 is to determine the three-dimensional structure of a human TCR bound to the HLA-DR2/MBP peptide complex, and to compare this structure to a complex with a crossreactive microbial peptide instead of the MBP peptide. The structural mechanisms of TCR crossreactivity will be further probed in mutagenesis experiments based on the structural data. The goal of Aim 2 is to define the extent and in vivo significance of TCR crossreactivity. The total number of peptides that can be recognized by MBP-specific T cell clones will be defined with peptide libraries in which each bead carries a distinct sequence. The in vivo activity of microbial peptides and the corresponding microbial proteins will be tested in a humanized mouse model of MS. This model is based on expression of HLA-DR2 and the human TCR that is the subject of the structural studies. Definition of the structural and functional basis of TCR crossreactivity has important implications for understanding the pathogenesis of MS and other chronic inflammatory diseases.